Despite the fact that current multi-modality treatment of human lymphomas can induce a high rate of complete remission, the majority of patients ultimately relapse and cannot be cured. The challenge, then, is to develop more effective strategies, with new immunotherapy modalities representing a promising area. Immunological tolerance to tumors and the lack of defined tumor antigens are the two major hurdles for the development of effective cancer immunotherapy. Dendritic cells (DCs) are the most potent antigen presenting cells. Recent compelling data suggest that DCs and their interaction with regulatory T cells (Treg) play critical roles in the generation and maintenance of immunologic self-tolerance. CD4+CD25+ Tregs are crucial for the prevention of autoimmune diseases. Rapidly accumulating data, including our preliminary data, provide evidence for an increased prevalence of CD4+CD25+ Treg cells in the tumor microenvironment and in the peripheral blood of patients with invasive cancers, indicating a potential negative role by these cells in tumor immunosurveillance. This proposal focuses on lymphoma immunotherapy and will address the two hurdles, and presents a plan to develop strategies to overcome these hurdles. The first hypothesis is that DCs are present in situ in the tumor, but are maintained in an inactive state either by regulatory T cells (CD4+CD25+) or by tumor cells, thereby leading to immunological tolerance to tumor antigens. Therefore effective immune responses to lymphomas do not occur naturally in patients with the tumors. The second hypothesis is that tumor antigens released by dying tumor cells after chemo- or radiation therapy are not efficiently cross-presented in vivo by DCs, due to either DC dysfunction or limited DC number in situ. Therefore systemic immune responses to lymphomas are not efficiently induced after chemo- or radiation therapy in patients with lymphomas. The major focus of this proposal is to characterize CD4+CD25+ regulatory T cells in human lymphomas, and to develop strategies in mouse models to counteract Treg cells (Pull) before delivering ex vivo expanded DCs to tumor sites (Push) after chemotherapy or local radiation therapy. The goal is to enhance cross-presentation of potential tumor antigens in vivo (Pull-Push strategy) and thus to induce systemic anti-tumor immunity and to eliminate the residual tumor. Hopefully these studies will pave the way for novel immunotherapy strategies which will be tested in future human clinical trials.